Open-Cell Foam Based Pathogen Remediation

ABSTRACT

An open-cell foam structure that is impregnated with a disinfectant and used to remove pathogens from air, water, and surfaces, and kill the pathogens

CROSS-REFERENCE TO RELATED APPLICATIONS

This application claims priority of Provisional Patent Application62/510,091, filed on May 23, 2017, the entire disclosure of which isincorporated herein by reference. This application is also acontinuation in part of and claims priority of both application Ser. No.15/987,664, filed on May 23, 2018, which is itself a continuation inpart and claims priority of application Ser. No. 15/454,626, filed Mar.9, 2017. The entire disclosures of each of these prior Applications areincorporated herein by reference.

BACKGROUND

Microbiological or biological contamination includes but is not limitedto pathogens, which include bacteria, fungi, mold, protozoa, virus, orother microorganism that can cause disease (collectively referred to aspathogens) and/or their associated toxins and byproducts. Suchbiological contamination is never in equilibrium or evenly distributedin water or air or on surfaces, let alone water or air that isconstantly flowing with many other variables to consider. Furthermore,to this day, there is not much known about biofilms and their respectiveformation and variability with changing conditions in aginginfrastructures and variability in water treatment methods.Instantaneous/grab sampling reflects what is in the water or air for asplit second, assumes the water or air being tested is in equilibrium,and does not take into consideration conditions like the mixture offresh water to bacteria of concern when the grab sample is taken.

The significance of bacteria, viruses and other pathogens of concern inour water and air and on surfaces are of increasing interest due totheir known and unknown effects on human health, antibiotic resistance,as well as the health of animals and plants, and effects on theecosystem. Animals and humans that are exposed to contaminated water orair can be exposed to pathogens.

Traditional sampling by collecting and analyzing a split second “grabsample” has several limitations. Among those limitations is theinability to detect transient biological contaminants or pathogens thatare discharging (including but not limited to releasing from biofilms)sporadically and diffusing through the water, water column, waterstream, or body of water on an irregular basis, and the limited samplesize that may contain only undetectable amounts of contaminants that arepresent at low concentration. Also, grab samples, by their nature, areinstantaneous and testing results only represent the volume of waterthat is in the sample bottle/container. As a fish does not swim in thewater for a split second and neither does a child, it is desired to havea sampling process that involves actual exposure over time and/orexposure to larger volumes of water beyond just the volume of thebottle/container (with a small volume of water from 250 ml to 1 liter)with corresponding identification of biological contaminants over timein the same way that life forms are exposed to contaminants over timethroughout more than just a limited volume of water. In essence, thissubject disclosure is based on biomimicry.

To better understand this disclosure, it is helpful to understand thebackground of biological contamination and pathogens more generally, andvarious methods that have been used to monitor biological contaminationand pathogens more generally. Heterotrophs are broadly defined asmicroorganisms that require organic carbon for growth. They includebacteria, yeasts and molds. Also, this includes bacteria that utilizeiron, copper, and phosphorus-related compounds as nutrients or foodsources. A variety of simple culture-based tests that are intended torecover a wide range of microorganisms from water are collectivelyreferred to as “heterotrophic plate count” or “HPC test” and “aerobicplate count” or “APC test” procedures. For purposes of this disclosureHPC and APC tests are used synonomously.

However, the terms “heterotroph” and “HPC” are not synonymous. There isno universal “HPC measurement.” Although standardized methods have beenformalized, HPC test methods involve a wide variety of test conditionsthat lead to a wide range of quantitative and qualitative results.Temperatures employed range from around 20° C. to 40° C., incubationtimes from a few hours to seven days or a few weeks, and nutrientconditions from low to high. The test itself does not specify theorganisms that are detected. Only a small proportion of themetabolically active microorganisms present in a water sample may growand be detected under any given set of HPC test conditions, and thepopulation recovered will differ significantly according to the methodused. The actual organisms recovered in HPC testing can also vary widelybetween locations, between seasons and between consecutive samples at asingle location.

Microorganisms recovered through HPC tests generally include those thatare part of the natural (typically non-hazardous) microbiota of water;in some instances, they may also include organisms derived from diversepollutant sources.

Microorganisms will normally grow in water and on surfaces in contactwith water as biofilms. Growth following drinking-water treatment isnormally referred to as “regrowth.” Growth is typically reflected inhigher HPC values measured in water samples. Elevated HPC levels occurespecially in stagnant parts of piped distribution systems, in domesticplumbing, in bottled water and in plumbed-in devices, such as softeners,carbon filters and vending machines. The principal determinants ofregrowth are temperature, availability of nutrients and lack of residualdisinfectant. Nutrients may be derived from the water body and/ormaterials in contact with water.

There is no evidence, either from epidemiological studies or fromcorrelation with occurrence of waterborne pathogens, that HPC valuesalone directly relate to health risk. They are therefore unsuitable forpublic health target setting or as sole justification for issuing “boilwater” advisories. Abrupt increases in HPC levels might sometimesconcurrently be associated with faecal contamination; tests for E. colior other faecal-specific indicators and other information are essentialfor determining whether a health risk exists. There is an unmet need forcost effective and efficient identification of biological contaminationin conjunction with HPC values; this is one of the benefits of thesubject disclosure.

In piped distribution systems, HPC measurements are assumed to respondprimarily to (and therefore provide a general indication of)distribution system conditions. These arise from stagnation, loss ofresidual disinfectant, high levels of assimilable organic carbon in thewater, higher water temperature, and availability of particularnutrients. In systems treated by chloramination or that contain ammoniain source waters, measurement of a variety of parameters, including HPC,but especially including nitrate and nitrite (which are regulated forhealth protection), can sometimes indicate the possible onset ofnitrification. This illustrates the importance of monitoring forexposure over time with the subject disclosure.

Some epidemiological studies have been conducted into the relationshipbetween HPC exposures from drinking water and human health effects.Other studies relevant to this issue include case studies, especially inclinical situations, and compromised animal challenge studies usingheterotrophic bacteria obtained from drinking-water distributionsystems. The available body of evidence supports the conclusion that, inthe absence of faecal contamination, there is no direct relationshipbetween HPC values in ingested water and human health effects in thepopulation at large. This conclusion is also supported indirectly byevidence from exposures to HPC in foodstuffs, where there is no evidencefor a health effects link in the absence of pathogen contamination.

There are opportunistic pathogens that may regrow in water but that arenot detected in HPC measurements, including strains of Legionella andnon-tuberculous mycobacteria. The public health significance ofinhalation exposure to some legionellae has been demonstrated. Again,since the HPC or APC is one general indicator, this is another exampleof why the subject disclosure is important with its ability to identifypathogenic bacteria including exposure over time.

The growth of bacteria in water distribution systems and water treatmentdevices has been recognized for many years. Such growth is affected bymany different factors, including the types of bacteria present in waterreleased from a water treatment plant, the temperature, disinfectantconcentration, the presence of sediment in the pipe work, the types andamount of nutrients present, and the rate of flow of the water. Many ofthese factors cannot be controlled, and thus microbial regrowth willcontinue to be investigated. The organisms involved in microbialregrowth are those that have been released from the water treatmentplant or that have been introduced into the distribution system at somepoint downstream of the water treatment plant. If it is assumed that thewater treatment plant is performing adequately, then the numbers ofbacterial pathogens released into the water distribution system will below, and those that are present are likely to be killed during transportin systems where residual disinfectant is present. However, a break inthe integrity of the distribution system (e.g., burst water main) canlead to the ingress of contaminated water. Such water may containorganisms that are potentially pathogenic for humans.

Many bacteria that enter the water distribution system are unable tosurvive or indeed colonize the distribution system, but many bacteria,including indicator bacteria such as Enterobacter, Citrobacter andKlebsiella, as well as potentially opportunistic pathogens such asAeromonas, Pseudomonas, Flavobacterium and Acinetobacter, are oftenfound in colonized water distribution systems.

Biofilms represent a specific form of bacterial colonization of waterdistribution systems. These specific forms determine the biostability ofthe microbial communities, their persistence and the release ofplanktonic cell microorganisms into the running water. The biofilmsinteract with waterborne pathogens and affect their persistence. Thepersistence of these pathogens is considerably increased if they form anew biofilm or colonize an existing one. The biofilms thus representbioreactors within water distribution systems, in which the resistanceof the microorganisms to disinfection is significantly increased. Thepotential for biofilm formation and growth is particularly high innarrow-gauge household plumbing. The colony count is directly correlatedwith the water volume that flows through these end-of-line systems.

SUMMARY

It is desirable to have an accurate and cost efficient method toremediate, collect and analyze water and air samples and surfaces forbiological contamination for large volumes of water and/or exposure overtime. It is also desirable to remediate pathogens in the air, in water,and on surfaces, using chemical-based and/or hydronium-baseddisinfectants.

This disclosure relates to remediating, detecting, removing, and killingbacteria, viruses, and other biological contaminants and pathogens fromwater and/or air and/or surfaces. Open-cell foam matrixcumulative/exposure testing not only identifies bacteria of concern andcorresponding colony formation units (“CFU”) but what the actualexposure is in the water or air or surfaces over time. The disclosurealso results in removal/filtration of bacteria, mold, viruses and otherorganisms and pathogens from the water or air or surfaces. The foam thatis used in the open-cell foam structures can be impregnated with one ormore biocides and/or disinfectants, or another chemical or substancethat can kill or neutralize pathogens which include bacteria, viruses,or other microbiological or biological organisms and other biologicalcontaminants.

One subject of this disclosure is an open-cell foam. The open-cell foamcan be made from various polymers. In one non-limiting example, the foamis produced from a copolymer of ethylene and alkyl acrylate. The foamcan comprise an elastomeric polyolefin. Examples of elastomericpolyolefins include but are not limited to ethylene methyl acrylate(EMA) and a single site initiated polyolefin elastomer (e.g. Dow orDuPont Dow Engage 8452) The open-cell foam is composed of a polyolefinelastomer which includes but is not limited relatively amorphouselastomers and/or includes blends of other polymers. The open-cellstructure of the various foams behaves as the alveoli of the human lungsin that it maximizes surface area which maximizes the efficacy of theopen-celled foam's ability to attract biological and relatedcontamination at the molecular level. The foam is hydrophobic and sorepels water. Because the foam is hydrophobic it does not promotebacterial or pathogen growth which makes it naturally antimicrobialwithout additional additives.

The open-celled foam structure provides high surface area due to theinterconnected structure of the individual cells. The oleophilic natureof the constituent polymer(s) prevents the absorption of water andpromotes absorption and adsorption of oils and other organic substancessuch as pathogens and viruses.

The structure that includes or is made of open-cell foam can befabricated from a very specific formulation for an open-cell foam.Specifically, this foam is produced from 80-100% ethylene acrylatecopolymer. Blends of LDPE (low density polyethylene) can be used also.One embodiment/formulation of this open-cell foam is described in U.S.Pat. No. 8,853,289, the disclosure of which is incorporated herein byreference. Another embodiment/formulation of this open-cell foam isdescribed in patent application US2013/0240451 A1, the disclosure ofwhich is incorporated herein by reference. While 80-100% EMA (ethylenemethyl acrylate) is one formulation of the open-cell foam that issubstantially non-polar, what is contemplated herein includes anyopen-cell foam produced from one or more polymers including but notlimited to EVA (ethyl vinyl acetate), EPDM (ethylene propylene dienemonomer), elastomers, LDPE, polypropylene, neoprene, styrene butadienerubber, ionic co-polymers, natural rubber, single site initiated ormetallocene polyolefins and equivalents. The preferred foam density isin the range of from about 1.0 pcf (pounds per cubic foot) to about 50.0pcf, but the foam can be any density less than the specific gravity ofwater (62.3 pcf at 70° F.). The open-cell foam can be extruded orproduced in a bun/batch process. The open-cell foam can be crosslinkedor non-crosslinked. Also, the open-cell foam can utilize either physicalblowing agents or chemical blowing agents. Furthermore, a bio-degradableinitiator may be added to the foam so that after use it will degradeover time in a landfill environment when disposed.

While open-cell polyurethane is one preferred material for the open-cellfoam discussed herein, what is contemplated herein includes anyopen-cell foam (with at least some of the cells open), and produced fromone or more polymers, such polymers including but not limited to EMA,ethylene vinyl acetate (EVA), ethylene-ethyl acrylate (EEA),ethylene-butyl acrylate (EBA), ethylene propylene diene monomer (EPDM),elastomers, polyolefin elastomers, low density polyethylene (LDPE),linear low density polyethylene (LLDPE), high density polyethylene(HDPE), polypropylene (PP), neoprene, styrene butadiene rubber, ionicco-polymers, other synthetic rubbers, natural rubber, chlorinatedpolyethylene (CPE), olefin block copolymers, ethylene maleic anhydridecopolymer, very low density polyethylene (VLDPE), singe site initiatedpolyolefins, metallocene catalyzed polyolefins, silane-modified polymers(including but not limited to silane grafted, silane functionalized, andsilane cross-linked polymers), maleic anhydride grafted polymers,styrene-butadiene-styrene copolymers, polyisoprene, and equivalents toany and all of these polymers. Silane modification of polymers can occurduring the manufacturing process of the open-cell foam, or as a separatestep after the foaming process, e.g., the silane can be applied inliquid form post-foaming. Further, specific silane-modified polymers maybe tailored to target specific contaminants that may be present in waterwith bacteria, such as VOCs and SVOCs related to oils and/or industrialchemicals, metals (e.g. copper, iron, etc.) or metalloids (e.g.phosphorus) and other petroleum products, and surfactants, including butnot limited to methylene blue active substances (MBAS).

The structure that includes or is made of open-cell foam can befabricated into a number of different forms to suit the particular usescenario or application. One structure is an assembly of strips,typically 0.5-0.75 inch×0.5-0.75 inch×12-18 inches. The strips arefastened together tightly at the center to form a structure withmultiple “fingers” or “blades.” This structure exposes a large surfacearea to the environment and allows flow through (between the fingers of)the indicator. In some examples, these structures are then fastened to arope line or similar tether with a weight at one end, and are submergedinto the water body, leaving foam structures at various depths. Anotheralternative is to have strips of the foam that are anchored to thebottom and extend to the surface, over the entire water column; this iscalled “eelgrass” since it looks like eelgrass that grows in the ocean.Other forms can include strips and smaller cubes and pieces in othershapes. Another form includes a design that is shaped like a “water bug”and is cast into the water or water column with a fishing rod. Any formcan be placed anywhere in the water column. Smaller pieces can be heldin place in nets or other containers, such as plexiglass. Other formsinclude placing the open-cell foam into a jar and effectively “swabbing”the water that is placed or run from a tap into the jar. Additionalforms include placing the open-cell foam structure into the air or at anair vent, waste water discharge location, a cooling tower, bathtub,shower, shellfish bed, river, lake, stream, and body of water, etc. andexposing to water or air for a period of time. The time period isnon-limiting and can be from a few seconds or minutes to hours or daysor more.

The monitoring/removal structure that comprises open-cell foam can alsobe made into a bracelet for humans to wear, in order to monitor aperson's exposure over time to biological contaminants.

The structure that includes or is made of open-cell foam can be designedto monitor the water or air or surfaces for biological contaminants. Thestructure that includes or is made of open-cell foam can also be used toremove the contaminants from the water or air or surfaces. Thestructures can be in the forms of eelgrass, cubes, small pieces, and/orstrips, and can be but need not be contained in a cylinder or net. Theseforms can be floating on the surface or suspended and/or submerged inthe water column using anchors.

The structure that includes or is made of open-cell foam can be wiped orpushed over a surface to be remediated from pathogens, a process that issometimes called “swabbing.” Swabbing can pick up pathogens andbiological contaminants that are on the surfaces that are swabbed.Swabbing can also pick up related contaminants. The foam material can beimpregnated with a chemical-based or hydronium-baseddisinfectant/biocide before or after it is exposed to pathogens.

The exposure time can be minutes, hours, days, weeks, or months,depending on the situation and desired results. The biological andrelated contaminants, pathogens and viruses are detected and removed andkilled by the impregnated open-cell foam structure. The open-cell foamcan be impregnated or infused with the disinfectant/biocide before orafter exposure to water, air, and/or surfaces. When used as anindicator, the un-impregnated structure can then be removed and testedfor the presence (and potentially the concentration) of contaminants.The structure that includes or is made from open-cell foam, includingbut not limited to fabricated in the form of eelgrass, can be used toabsorb and concentrate pathogens, thereby removing pathogens from waterinto the open-cell foam capillary network.

In some examples (e.g., in shellfish beds where the shellfish cannot beexposed to the disinfectant), the structure that includes or is madefrom open-cell foam is located in the bed, where it picks up pathogens.It is then removed from the water, and the disinfectant is then infusedor applied to the open-cell foam after the open-cell foam is removedfrom the water, to kill the pathogens. If desired, the water,disinfectant and killed pathogens can be removed from the foam bywringing or otherwise squeezing the foam. The foam can then be re-usedto pick up and remove more pathogens. This process of exposure, thendisinfectant application, then squeezing, and re-use can be applied towater or air or surfaces.

In some examples biochar is used together with the foam. Biochar is anexcellent adsorbent of organic matter, including pathogens. Biochar canbe incorporated directly into the foam and/or it can be incorporated ina porous structure (like a sock) that is coupled to or carried by thestructure that includes or is made up of the foam. Biochars and its usedtogether with open-cell foam are further described in U.S. PatentApplication Publication 2020/0156960, the entire disclosure of which isincorporated herein by reference.

Since the structure that includes or is made from open-cell foam canspan different depths (or heights in the air), the results can determinethe presence (and concentration) of one or more biological contaminantsat different depths of the water (or air) or water column, from thesurface to the bottom, or from the ground to a desired height, forexample, and as desired.

Advantages of this biological remediator/indicator are its efficientcost, ease of deployment, durability during deployment and in use, andability to collect samples for analysis over large volumes of water overan extended time period.

Upon retrieval of the indicator, the open-cell foam can be placed into asealed container and sent to a qualified lab to test the open-cell foammatrix with various testing methods. More detail is provided elsewherein this document.

Another significant attribute of the open-cell foam in one embodiment isutilizing a non-polar polymer with the result that the foam has no ioniccharge. During the recent pandemic of 2020, the fact that paper towels,cotton rags, and cellulose sponges deactivate disinfectant because oftheir negative ionic charges came to the forefront as live pathogensincluding a virus like SARS-CoV-2 was left on surfaces therebytransmitting COVID-19 and infecting humans which include front linehealth care workers who were wearing full PPE gear and N95 masks. Papertowels, cotton rags, and cellulose sponges all have a negative ioniccharge where the majority of disinfectants used (e.g. quats) have apositive ionic charge thereby cancelling each other out and renderingthe disinfectant inactive leaving live pathogens on surfaces.

Deactivation of disinfectant is sometimes referred to as “quat binding.”The phenomenon of quat binding occurs when the active ingredient(quaternary ammonium chloride) becomes attracted to and absorbed intofabrics. The science behind how this happens is simple: Quats arepositively charged ions and cotton and other natural textiles arenegatively charged; positive attracts negative.

In United States Patent Publication US 2007/0142261 A1 entitled, “WIPERFOR USE WITH DISINFECTANTS” (incorporated by reference herein), theproblems with quat binding and deactivation of disinfectant are wellestablished.

What makes the subject open-cell foam invention unique is the highsurface area for absorption and release of active disinfectant thatsubstantially mitigates the risk of gaps in surface coverage or notmaking contact with live pathogens on surfaces. The high surface areacombined with a neutral ionic charge, assures no deactivation ofdisinfectant/biocide along with maximum delivery of disinfectant/biocideper square inch of surface area.

Furthermore, based on validation testing, these results have proven theability of the open-cell foam biological indicator to detect Legionellaat low levels where conventional grab samples can show non-detects whenin fact Legionella was present. Data is set forth elsewhere.

In one aspect a method of killing pathogens includes providing astructure that comprises an open-cell polymer foam, impregnating theopen-cell foam with a substance that comprises a biocide or anotherchemical that can kill pathogens, and exposing the structure to thepathogens such that the impregnated open-cell foam contacts and killsthe pathogens.

In an example the pathogens are on a surface or in air or in water. Inan example exposing the structure to the pathogens comprises moving thestructure over the surface such that the impregnated open-cell foamcontacts the pathogens, removes the pathogens from the surface, andkills the pathogens.

In another aspect a method of remediating pathogens that are on asurface includes providing a structure that comprises an open-cellpolymer foam, applying a disinfectant that can kill pathogens to one orboth of the surface and the open-cell polymer foam, and moving thestructure over the surface such that the open-cell foam contacts thepathogens, removes the pathogens from the surface, and the pathogens arekilled. In an example the disinfectant comprises at least one of achemical-based disinfectant and a hydronium-based disinfectant.

In another aspect a method of remediating pathogens includes providing astructure that comprises an open-cell polymer foam, exposing thestructure to at least one of air and water such that the open-cell foamcontacts pathogens, and either before or after the exposing step,applying to the open-cell polymer foam a disinfectant that can killpathogens, to kill pathogens that come into contact with the foam.

In an example the disinfectant comprises at least one of achemical-based disinfectant and a hydronium-based disinfectant. In anexample the structure comprises a face mask that is configured to beworn by a person covering at least one of the mouth and nostrils. In anexample the structure is located at least in part in a buildingventilation system. In an example the disinfectant is applied after theexposing step. In some examples the method further includes wringing outthe foam after the application of the disinfectant, to remove at leastsome of the disinfectant from the foam. In an example the method furtherincludes reusing the wrung-out foam by exposing it a second time to atleast one of air and water such that the open-cell foam contactspathogens.

In an example the application of disinfectant comprises impregnating thefoam with the disinfectant. In some examples the method further includesincorporating biochar into the open-cell foam or coupling biochar to thestructure. In an example the biochar is held in a porous container thatis coupled to the structure.

BRIEF DESCRIPTION OF THE DRAWING

The drawing depicts one non-limiting example of the placement ofopen-cell foam material into a body of water, as a step in thecollection and removal from the water and neutralization or killing ofbiological and microbiological contaminants and pathogens.

DETAILED DESCRIPTION OF EXAMPLES

Methods of removing biological contaminants and pathogens from a body ofwater or the air or surfaces are disclosed. The methods also generallyinvolve killing the pathogens using a disinfectant/biocide that iscarried by the open-cell foam structure. Since the open-cell foam has avery large surface are, more disinfectant can be carried as compared toother wipes. Also, the oleophilic/hydrophobic nature of the open-cellfoam is effective to absorb/adsorb large amounts of pathogens, leadingto better remediation results as compared to the use of paper towels orother wipes or wipers.

As a first step, an open-cell foam material (or other foam materials, asdescribed elsewhere herein) can be placed into water or into the air, orwater or air can be passed though the material. The placement can be atone or more locations in the body of water or air, and at one or moredepths or heights in the body of water or in the air. After desiredexposure times, one or more separate portions of the open-cell foammaterial are removed from the water or air. The foam can be impregnatedwith a disinfectant/biocide either before or after it picks up thepathogens from the air, water and/or surfaces it is moved over. In someexamples the presence in the removed separate portions of one or morebiological contaminants that were removed from the water or air by theopen-cell foam material are then determined, typically by standardtesting procedures well known in the art for the particular type ofbiological contaminant(s).

In an example the foam structure is placed in a building ventilationsystem such that ventialation air passes through the foam. The foamcaptures pathogens in the air. The foam can be treated with the liquiddisinfectant either before or after it is exposed to the air, to killthe pathogens. The previously-exposed foam can be wrung out or squeezedto remove liquid disinfectant, and then reused for additional exposuresand pathogen remediation using additional disinfectant. In anotherexample the foam can be configured as a face mask that is worn by aperson covering one or both of the mouth and nostrils. The foam absorbspathogens such as virus particles. The pathogens can then if desired bekilled by applying the disinfectant to the foam, or the foam can besafely disposed of without the use of disinfectant. If foam is to bere-used, used disinfectant can be removed, e.g., by wringing out orsqueezing the foam to remove liquid disinfectant. The foam can then beused all over again.

There are several different preferred water testing methods with theopen-cell foam. Non-limiting examples include the following. In a firstexample, a grab sample can be taken by placing a piece of the open-cellfoam in a sample jar and then partially or fully filling the jar withwater. The foam can be removed for testing after any desired exposuretime. If necessary to help preserve specimens that are collected by thefoam, the container with water and foam can be placed on ice until thefoam is ready to be tested; however, ice is not necessarily required. Ina second example, the open-cell foam can be placed directly into astream or body of water to be tested. Exposure times can vary;non-limiting examples are 5, 10, or 20 minutes. The foam is then removedfrom the water and tested. In a third example, cumulative testing can beaccomplished by placing the foam into water to be tested, and thenperiodically removing portions of the foam at different exposure times.

The methods are effective both to determine the presence of and killpathogens and biological contaminants in the water or air or onsurfaces, and also to remove such contaminants from the water or air orsurfaces. The methods thus can be used for contaminant detection and/orfiltration or remediation.

The drawing depicts three groups of strips or “blades” of open-cell foammaterial 12, 14 and 16. Each group has multiple strips that are heldtogether at about their centers. The groups are fastened to a line 32that is held on the bottom 24 of water body 20 by weight or anchor 30.In this example group 16 floats on the water surface 22, while groups 12and 14 are held at different depths below the surface. This disclosureallows for the placement of open-cell foam material at any one or moreheights of a body of water and/or the air, and at one or more locationsin the body of water or air. Various non-limiting methods of exposingthe open-cell material to water or air are described herein; any suchmethod can be used as desired or as necessary depending on the body ofwater or the air mass, and/or the testing regime that is desired underthe circumstances.

After desired exposure times, one or more portions of the foam materialare removed from the water or air. This can be done by clipping orcutting a piece of foam, or removing an entire group or other portion orseparate piece of foam, for example. The exposure times can be fromseconds to minutes to hours to days to weeks to months, depending on theparticular testing regime. Since the open-cell foam absorbs and adsorbsbiological contaminants, the removed portions of the foam can be testedfor particular biological contaminant(s) that are expected or are beinginvestigated. The foam can act as an accumulator for these biologicalcontaminants. Also, the different locations and different exposure timesallow for a tailored review of biological contaminants, their locations,and their movement within the water or air.

The subject materials have been used in testing of potable water. Testmethods and results follow.

Results of uses of the biological indicator in water are disclosed inthe appendices 1-5 of the priority Provisional application, which areincorporated by reference herein in their entireties. A brief discussionof those appendices follows.

Appendix 1 that was part of the Provisional Application that isincorporated herein by reference (four pages) is a report from anindependent testing laboratory that details the study design,procedures, and results, for comparison of grab samples (prior art) totesting using the open-cell foam of the present disclosure in potablewater. The results prove that the open-cell foam acts as a biologicalindicator, as it is effective to remove and detect Legionella at lowlevels, where conventional grab samples can show non-detects when infact Legionella is present.

Appendix 1 included the following:

A purpose of this study was to identify an effective method for theextraction of Legionella from an open-cell foam environmental indicatorsampling device. Replicate sponge devices (i.e., pieces of the open-cellfoam) were indirectly inoculated with a mixed suspension of freshLegionella cultures at three target concentrations: low (1-10 CFU/mL),medium (10-100 CFU/mL) and high (100-1,000 CFU/mL). The recovery anddetection procedure of the pathogen was evaluated using a non-ionicsurfactant (Polysorbate 80) in conjunction with a maceration extractionprocess and nutritive media (BCYE agars) culturing following amodification of the Centers for Disease Control and Prevention (CDC)“Procedures for the Recovery of Legionella from the Environment”,January 2005. A summary of the study design is presented in Table Abelow.

TABLE A Legionella Recovery Study Design Summary Maceration MixedLegionella Target Target Extraction suspension Matrix LevelConcentration Procedure Surfactant Legionella pneumophila Sterile TapLow 1-10 CFU/mL Blending Polysorbate ATCC¹ 33152 Water 80³ Legionelladumoffii Medium 10-100 CFU/mL QL14012²-1A High 100-1,000 CFU/mLLegionella micdadei QL145022-1A ¹ATCC: American Type Culture Collection²QL: Q Laboratories, Inc. Culture Collection ³The polysorbate wasTween ™ 80, which is a registered trademark of Croda Americas, Inc.

The study included three replicate open-cell sponge samples indirectlyinoculated for each target contamination level with Legionella species.For each contamination level, one liter of sterile tap water wasinoculated using a mixed suspension of the Legionella cultures that hadbeen diluted to the targeted levels. To simulate real-worldenvironmental sampling, each open-cell device was submerged and allowedto absorb the contaminated water for 3-5 minutes. During submersion, thesponges were mixed in a bobbing motion using sterile pipettes. Thesponges were then placed into the original sample glass vial andapproximately 200 mL of the contaminated water added and the lid tightlycapped. Samples remained at ambient temperature (20-24° C.) forapproximately 24 hours prior to analysis.

Legionella Extraction and Detection Extraction

All metals rings and zip ties were aseptically removed from each spongesample prior to transferring all sample contents to a sterile laboratoryblender jar. A one milliliter volume of a sterile, non-ionic surfactant,Tween™ 80, was added to each blender jar to facilitate the release ofany Legionella organisms that may be present within the pores of thesampling device.

Open-cell sponge samples were blended for two minutes and the jarsallowed to rest for approximately ten minutes, which provided sufficienttime for the sponge particulate matter to float to the surface. Theliquid portion of each blender jar was aseptically transferred tosterile conical tubes and centrifuged at 5500×g for thirty minutes atambient temperature (20-24° C.). All but five milliliters of thesupernatant was aseptically removed and discarded into approvedbiohazard containers.

Detection

The remaining five milliliters of sample was homogenized by vortex andan aliquot spread plated onto BCYE, PCV, GPCV and PCV (−)microbiological agar plates and incubated aerobically at 35±1° C. toencourage the proliferation of Legionella organisms. The presence orabsence of typical Legionella colonies based on morphology and/orfluorescence was determined after 72 to 96 hours of incubation. If anyagar plates did not appear to contain typical colonies, incubation wasextended for an additional seven days.

Typical colonies from each contamination level replicate were re-struckto selective and non-selective media. Typical colonies were thenconfirmed via serological latex agglutination and molecularidentification using the Bruker MS Biotyper.

The results obtained from this method development study indicate thatoverall, the extraction procedure had positive outcomes for removingLegionella microorganisms the open-cell foam environmental samplingdevice. The novel open-cell foam sponges evaluated in this study wereinoculated at levels as low as about 4 (e.g. 3.5) CFU/mL, or as high asapproximately 250 CFU/mL. Inoculation of the device paralleled actualsampling procedures employed in the field. Whether the pathogen ispresent at a level of a few cells or many thousands of cells permilliliter, the ability to capture, extract, and detect the organismreliably and consistently is paramount to maintaining the good health ofthe building occupants. The detection of Legionella is dependent uponthe sampling device or procedure used in addition to the laboratorymethod employed. One cannot be successful without the other.

The cultural detection and confirmation of Legionella at all levels forall replicates demonstrates the method has applicability as a viableoption for Legionella analysis in routine water samples. See Tables Band C for detailed inoculum and recovery results.

TABLE B Inoculum Results Mixed Legionella Mixed Inoculum Extractionsuspension Matrix Target Level Concentration Procedure SurfactantLegionella pneumophila Sterile Tap Low 3.5 CFU/mL Blending PolysorbateATCC 33152 Water 80 Legionella dumoffii Medium 20.6 CFU/mL QL14012-1AHigh 247.5 CFU/mL Legionella micdadei QL145022-1A

TABLE C Detailed Recovery Results Confirmation Examination for TypicalLegionella Slide Agglutination Bruker Contamination PCV GPCV PCV PCVTest Biotyper Level/Replicate BCYE A B A B (−) BCYE^(a) (−) SBA 1 2-15L. spp. Result ID Low A + + − + + − + + − − + + − − Positive Legionellapneumophila Low B + + + − + − + + − − + − + Positive Legionellapneumophila, Legionella dumoffii Low C + + + + + − + + − − + + − −Positive Legionella pneumophila Medium A + + + + + − + + − − + − +Positive Legionella pneumophila, Legionella dumoffii Medium B + + + + +− + + − − + − + Positive Legionella pneumophila, Legionella micdadeiMedium C + + + + + − + + − − + + − − Positive Legionella pneumophilaHigh A + + + + + − + + − − + + − − Positive Legionella pneumophila HighB + + + + + − + + − − + + − − Positive Legionella pneumophila HighC + + + + + − + + − − + − + Positive Legionella pneumophila, Legionellamicdadei Sterility Control − − − − − − − − − NA NA NA Typical NANegative − − − − − − − − − − − − Typical NA Control PositiveControl + + + + + − + − − + − + Typical Legionella pneumophila ^(a)Twotypical Legionella colonies picked for serological confirmation andmolecular identification

The procedure to extract Legionella from the open-cell foamenvironmental sampling device was adapted based on previous works fordetecting Legionella from environmental samples. The positive outcomesof this study following the procedures presented above, as well asexperience working with similar sampling devices, has promptedpossibilities of streamlining the method to better suit the workflow ina routine laboratory environment. Blending the device requires sterilelaboratory blender jars with sharp blades and potentially poses a safetyrisk if not performed in a careful manner and in a Biological SafetyCabinet (BSC). One alternative to blending is to place the samplingdevice into a common sterile laboratory blender bag with Tween™ 80 andextract the bacteria by homogenizing with a laboratory paddle blender.This procedure would not only decrease the time required for processingthe sample but also allow for the use of readily available disposablesterile materials used by a majority of testing laboratories. Selectingblender bags would have the additional benefit of increasing the ease ofuse factor, thereby improving laboratory technician efficiency.

The inoculation method utilized in this laboratory study followed theprescribed, real world best practices for correctly sampling with thesponge device: the glass jars containing the sponges were filled withthe sample water to be tested.

Appendices 2-5 that were part of the Provisional Application that isincorporated herein by reference (two pages each) included reports fromindependent lab testing from Flint Mich.—where bacteria has been acontinued challenge with the water distribution system and potentialreported human health effects. For the testing with the open-cell foambiological indicator the lab used the following tests and methods:

TEST METHOD Aerobic Plate Count (APC) FDA Bacteriological AnalyticalManual Coliform AOAC 991.14 Legionella Centers for Disease Control(January 2005) Microbial Identification Bruker MALDI Biotyper (Q LabsSOP #10- MIDL-METH-001A)For the testing for the water grab samples the lab used the followingtest and method:

TEST METHOD Aerobic Plate Count (APC) Standard Methods for theExamination of Water and Wastewater, 22^(nd) Edition

Appendix 2 shows lower to <10 or <1 (non-detect) APC counts on grabsamples while the open-cell foam biological indicator (“Waterbug”) showsAPCs in the millions and identifies bacteria of concern. The followingis from appendix 2.

The following results were obtained from the samples submitted forassay:

Methodology

TEST METHOD Aerobic Plate Count (APC) Standard Methods for theExamination of Water and Wastewater, 22^(nd) Edition

Results

Sample No. IDENTIFICATION OF SAMPLE APC/mL 1 1608640-01A 4,400 (UpstairsBath Grab for Bacteria/Fungi) 2 1608640-05A <10 (Water Meter Grab forBacteria/Fungi)

Methodology

TEST METHOD Aerobic Plate Count (APC) FDA Bacteriological AnalyticalManual Identification Gram Stain & VITEK

Results

Sample IDENTIFICATION No. OF SAMPLE APC/sponge Identification 11608640-03B 3,800,000 Acinetobacter (Upstairs Bath junii, WaterBug Grab)Brevibacillus 2 1608640-07B 2,900,000 Pseudomonas (Water Meteraeruginosa/ WaterBug Grab) Pseudomonas putida,

Appendix 3 shows no APC count on the grab sample while the open-cellfoam biological indicator (“Waterbug”) shows an APC count of >570,000and identifies bacteria of concern Pseudomonas aeruginosa.

The following results were obtained from the samples submitted forassay:

Methodology

TEST METHOD Aerobic Plate Count (APC) Standard Method for theExamination of Water and Wastewater, 22^(nd) Edition

Results

Sample No. IDENTIFICATION OF SAMPLE APC/mL 1 1609132-04A (Amber Grab forBacteria/Fungi) <1

Methodology

TEST METHOD Aerobic Plate Count (APC) FDA Bacteriological AnalyticalManual Identification Gram Stain & VITEK

Results

Sample IDENTIFICATION No. OF SAMPLE APC/sponge Identification 11609132-03B (WaterBug Grab >570,000 Pseudomonas 5 mins) aeruginosa

Appendix 4 shows lower to <1 (non-detect) APC counts on grab sampleswhile the open-cell foam biological indicator (“Waterbug”) showsAPCs>570,000 and identifies bacteria.

The following results were obtained from the samples submitted forassay:

Methodology

TEST METHOD Aerobic Plate Count (APC) FDA Bacteriological AnalyticalManual Identification Gram Stain & VITEK

Results

Sample IDENTIFICATION No. OF SAMPLE APC/mL Identification 1 1609134-01A(Water <1 N/A Meter Amber Grab for Bacteria/Fungi) 2 1609134-05A (Shower3,100 Bacillus simplex Amber Grab for Bacteria/Fungi)

Methodology

TEST METHOD Aerobic Plate Count (APC) FDA Bacteriological AnalyticalManual Identification Gram Stain & VITEK

Results

Sample IDENTIFICATION No. OF SAMPLE APC/sponge Identification 11609134-03B >570,000 Pseudomonas (Water Meter fluorescens Grab WaterBug)2 1609134-07B >570,000 Acinetobacter (Shower Grab species WaterBug)

Appendix 5 shows low APC counts (11,000 and <1) while the open-cell foambiological indicator (“Waterbug”) shows APC counts of 150,000 and>570,000.

The following results were obtained from the samples submitted forassay:

Methodology

TEST METHOD Aerobic Plate Count (APC) FDA Bacteriological AnalyticalManual Identification Gram Stain & VITEK

Results

Sample IDENTIFICATION No. OF SAMPLE APC/sponge Identification 11609133-03B 150,000 Delftia acidovorans (Murphy Water Meter WaterBugGrab) 2 1609133-07B >570,000 Brevundimonas (Murphy Showerdiminuta/vesicularis Grab WaterBug)

Methodology

TEST METHOD Aerobic Plate Count (APC) FDA Bacteriological AnalyticalManual Identification Gram Stain & VITEK

Results

Sample IDENTIFICATION No. OF SAMPLE APC/mL Identification 1 1609133-01A(Murphy Water 11,000 Rhodotorula sp. Meter Amber Grab forBacteria/Fungi) 2 1609133-05A (Murphy <1 N/A Shower Amber Grab forBacteria/Fungi)

A second report from an independent testing laboratory details the studydesign, procedures, and results, for the evaluation of the ability forsix different types of the subject open-cell foam sampling devices(i.e., WaterBugs) in recovering and releasing select bacteria from awater source. In this study, a bulk lot of sterile tap water wasinoculated with Pseudomonas aeruginosa. Traditional “grab samples”consisting of three (3) replicate 100 mL volumes were collected toestablish starting baseline bacterial counts for evaluation purposes.WaterBug sampling devices, comprised of six different designformulations, and in replicates of three, were submerged for a total of20 minutes. During submersion the inoculated water was periodicallymixed to maintain homogeneity and even distribution of the bacteria.After 20 minutes had elapsed, each WaterBug was transferred individuallyto a sterile stomacher bag. Customary laboratory procedures forextracting bacteria from matrices involve the use of a laboratory paddleblender, or “stomacher”. One point of focus for this study was todetermine the stomaching time for optimal recovery; therefore an aliquotfrom each bag was removed after being stomached for 30 seconds, 1minute, and 2 minutes. At each time point, the aliquot was diluted asappropriate and the concentration of target organism determined usingstandard microbiological plate count techniques. Final bacterial countsof the inoculated water were determined after the WaterBugs were removedby obtaining three 100 mL traditional grab samples and enumerating aspreviously described. A summary of the WaterBug formulations tested andstudy summary is presented in Table A below.

TABLE A Pseudomonas Retention and Release Study Design Summary PlatingWaterBug Target Extraction Medium/ Formulation Matrix Organism ProcedureIncubation A: Open-cell Sterile Pseudomonas Stomaching MacConkey EMA Tapaeruginosa (30 s, 1 min, agar 35° C. B: Closed-cell Water ATCC 15442 2min) for 24 ± EMA 2 hours C1: Open-cell LDPE/8452 C2: Open-cell EVA/8452Large-cell C3: Open-cell EVA/8452 Small-cell D: Open-cell urethanePseudomonas aeruginosa Extraction and Enumeration

Extraction

Prior to submersing the WaterBugs, 3×100 mL grab samples were taken fromthe inoculated sterile tap water. The WaterBugs were removed after 20minutes of submersion in the inoculated sterile tap water and werestomached for 30 seconds, 1 minute, and 2 minutes. An aliquot of steriletap water was removed at each time point. An additional 3×100 mL grabsamples were taken from the inoculated sterile tap water once theWaterBugs had been removed.

Enumeration

The grab samples and the aliquots of the inoculated sterile tap waterremoved at the three pre-determined time points for each of the WaterBugformulations was plated onto MacConkey agar in duplicate. The dilutionswere spread plated and incubated at 35±1° C. for 24±2 hours. Typicalcolonies were enumerated and recorded as CFU/plate, then averaged andmultiplied by the dilution factor to determine the amount ofmicroorganisms present in the inoculated sterile tap water sample at thebeginning and end of testing as well as the concentration recovered fromeach of the different sponge design formulations.

The average CFU/mL, expressed as normalized values (Log ₁₀), recoveredfrom each WaterBug design formulation was compared to the averageinitial grab samples prior to submersion to obtain percent recovery ateach time point in the bacterial extraction process (30 sec., 1 min., 2min.). Of the six WaterBug formulations tested, Type A: Open-cell EMAdemonstrated the highest retention and subsequent release of theinoculating organism at 88.8% after a 1 minute stomaching time period.Type B: Closed-cell EMA demonstrated the lowest retention and releaseafter 2 minutes of stomaching at 70.5%. Type C2: Open-cell EVA/8452Large-cell was the only formulation to show an increase in percentrecovery at the final stomaching time point. This may suggest that itperformed best at retaining liquid and bacteria compared to the otherformulations; however, the concentration of trapped bacteria that werereleased was less than other designs on average. Comparing thedifference of means between the initial grab sample counts and mean Log₁₀ counts for each sampling time point demonstrates significantdifferences (>0.5 Log ₁₀) with several of the design formulations.Tables B and C present the results of the percent recovery and thedifference of means.

TABLE B Grab Sample Recovery Results Grab Samples Average CFU/mL Log₁₀CFU/mL Initial 3.9 × 10² 2.5911 Final 3.3 × 10¹ 1.5185

TABLE C Sponge Formulation Statistical Data 30 sec. Stomach 1 min.Stomach Sponge Log₁₀ % Mean Log₁₀ % Formulation CFU/mL CFU/mL Recovery¹Difference² CFU/mL CFU/mL Recovery¹ A: Open-cell 2.0 × 10² 2.3010 88.80.2901 2.0 × 10² 2.3010 88.8 EMA B: Closed-cell 9.7 × 10¹ 1.9868 76.70.6043 1.0 × 10² 2.0000 77.2 EMA C1: Open-cell 1.3 × 10² 2.1139 81.60.4772 1.2 × 10² 2.0792 80.2 LDPE/8452 C2: Open-cell 1.4 × 10² 2.146182.8 0.4450 1.3 × 10² 2.1139 81.6 EVA/8452 Large-cell C3: Open-cell 1.0× 10² 2.0000 77.2 0.5911 1.1 × 10² 2.0414 78.8 EVA/8452 Small-cell D:Open-cell 1.6 × 10² 2.2041 85.1 0.3870 1.2 × 10² 2.0792 80.2 Urethane 1min. Stomach 2 min. Stomach Sponge Mean Log₁₀ % Mean FormulationDifference² CFU/mL CFU/mL Recovery¹ Difference² A: Open-cell 0.2901 1.7× 10² 2.2304 86.1 0.3607 EMA B: Closed-cell 0.5911 6.7 × 10¹ 1.8261 70.50.7650 EMA C1: Open-cell 0.5119 1.3 × 10² 2.1139 81.6 0.4772 LDPE/8452C2: Open-cell 0.4772 1.6 × 10² 2.2041 85.1 0.3870 EVA/8452 Large-cellC3: Open-cell 0.5497 1.0 × 10² 2.0000 77.2 0.5911 EVA/8452 Small-cell D:Open-cell 0.5119 1.1 × 10² 2.0414 78.8 0.5497 Urethane ¹% recoverycalculated using the Log₁₀ CFU/mL mean average at each sampling timepoint and the initial grab sample Log₁₀ CFU/mL mean average ²A meandifference absolute value of greater than 0.5 indicates a statisticalsignificant difference between counts

In some examples a chemical-based and/or hydronium-based disinfectant isused with the open-cell foam structure. Chemical-based disinfectantsinclude but are not limited to alcohols, hydrogen peroxide, quaternaryammonium chlorides (quats), and other pathogen disinfectants that arewell known in the field and so not further described herein, includingbut not limited to those described in U.S. Pat. Nos. 6,331,514 and8,940,792 and U.S. Patent Application Publication 2007/0142261.Hydronium-based disinfectants are well known in the field and so are notfurther described herein, including but are not limited to thosedescribed in U.S. Pat. Nos. 10,039,696 and 9,204,633. The disclosures ofeach of these patents and publications are incorporated by referenceherein for all purposes.

In an example a number of strips of open-cell foam that were infusedwith hydronium-based Hy-IQ sanitizer available from Aphex BiocleanseSystems, Inc. of Pittsford, N.Y. were placed into contaminated waterfrom a body of water contaminated with fecal coliform from awater-treatment plant. All pathogens were killed.

The invention is not limited by the above description but rather issupported by it. Other options will occur to those skilled in the artand are within the scope of the following claims.

What is claimed is:
 1. A method of killing pathogens, comprising:providing a structure that comprises an open-cell polymer foam;impregnating the open-cell foam with a substance that comprises abiocide or another chemical that can kill pathogens; and exposing thestructure to the pathogens such that the impregnated open-cell foamcontacts and kills the pathogens.
 2. The method of claim 1, wherein thepathogens are on a surface or in air or in water.
 3. The method of claim2, wherein exposing the structure to the pathogens comprises moving thestructure over the surface such that the impregnated open-cell foamcontacts the pathogens, removes the pathogens from the surface, andkills the pathogens.
 4. A method of remediating pathogens that are on asurface, comprising: providing a structure that comprises an open-cellpolymer foam; applying a disinfectant that can kill pathogens to one orboth of the surface and the open-cell polymer foam; and moving thestructure over the surface such that the open-cell foam contacts thepathogens, removes the pathogens from the surface, and the pathogens arekilled.
 5. The method of claim 4, wherein the disinfectant comprises atleast one of a chemical-based disinfectant and a hydronium-baseddisinfectant.
 6. A method of remediating pathogens, comprising:providing a structure that comprises an open-cell polymer foam; exposingthe structure to at least one of air and water such that the open-cellfoam contacts pathogens; and either before or after the exposing step,applying to the open-cell polymer foam a disinfectant that can killpathogens, to kill pathogens that come into contact with the foam. 7.The method of claim 6, wherein the disinfectant comprises at least oneof a chemical-based disinfectant and a hydronium-based disinfectant. 8.The method of claim 6, wherein the structure comprises a face mask thatis configured to be worn by a person covering at least one of the mouthand nostrils.
 9. The method of claim 6, wherein the structure is locatedat least in part in a building ventilation system.
 10. The method ofclaim 6, wherein the disinfectant is applied after the exposing step.11. The method of claim 10, further comprising wringing out the foamafter the application of the disinfectant, to remove at least some ofthe disinfectant from the foam.
 12. The method of claim 11, furthercomprising reusing the wrung-out foam by exposing it a second time to atleast one of air and water such that the open-cell foam contactspathogens.
 13. The method of claim 6, wherein the application ofdisinfectant comprises impregnating the foam with the disinfectant. 14.The method of claim 6, further comprising incorporating biochar into theopen-cell foam or coupling biochar to the structure.
 15. The method ofclaim 14, wherein the biochar is held in a porous container that iscoupled to the structure.